A Kunitz-type glycosylated elastase inhibitor with one disulfide bridge

A Kunitz-type glycosylated elastase inhibitor with one disulfide bridge

Author Sumikawa, J. T. Google Scholar
Nakahata, A. M. Google Scholar
Fritz, H. Google Scholar
Mentele, R. Google Scholar
Sampaio, M. U. Google Scholar
Oliva, MLV Google Scholar
Institution Universidade Federal de São Paulo (UNIFESP)
LMU
Max Planck Inst Biochem
Abstract A glycosylated Bauhinia rufa elastase inhibitor (gBrEI) was purified and characterized using acetone precipitation, affinity chromatography on concanavalin A-Sepharose, ion-exchange chromatography on a HiTrap Q column, size exclusion chromatography on a Superdex 200 column and reverse-phase chromatography on a C-18 column. gBrEI inhibited pancreatic porcine elastase with an equilibrium dissociation constant (K-i) of 6.18 x 10(-8) M, but it did not inhibit human neutrophil elastase, bovine trypsin, human plasma kallikrein or porcine pancreatic kallikrein. On SDS-electrophoresis, gBrEI appeared as a single 20-kDa band, also after reduction. Schiff reagent staining indicated a carbohydrate portion in the protein, which was confirmed by mass spectrometry. the glycosylated site was Asn(38), and a carbohydrate portion of 1.17 kDa was identified. gBrEI was found to contain 144 amino acid residues, and a FASTA database analysis showed that it belongs to the plant Kunitz-type inhibitor family. Val(66) was identified as reactive site P1 residue by comparison of conserved positions in the sequences. Since gBrEI harbors a single disulfide bridge, it may be considered a new type of Kunitz inhibitor, intermediate between the classical kunitz inhibitors, which contain two disulfide bridges, and those from B. bauhinioides, which do not have such bridges.
Keywords Bauhinia rufa
elastase/antagonists and inhibitors
glycoproteins
Kunitz inhibitor
leguminosae
plant
serine endopeptidases
Language English
Date 2006-04-01
Published in Planta Medica. Stuttgart: Georg Thieme Verlag Kg, v. 72, n. 5, p. 393-397, 2006.
ISSN 0032-0943 (Sherpa/Romeo, impact factor)
Publisher Georg Thieme Verlag Kg
Extent 393-397
Origin http://dx.doi.org/10.1055/s-2005-916237
Access rights Closed access
Type Article
Web of Science ID WOS:000237192200002
URI http://repositorio.unifesp.br/handle/11600/28827

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